Prof. Dr. Joachim Wittbrodt

Vertebrate eye and brain development and regeneration

The vertebrate eye is composed of neuroectodermal (optic cup) and surface ectodermal (lens, cornea) derivatives and it emerges from an epithelial Anlage by inductive interactions beginning al late gastrula stages. Under the influence of midline signaling during neurulation the single retina Anlage is split into two retinal primordia localized in the lateral wall of the forebrain.
Subsequent evagination of the primordia results in the formation of optic vesicles that differentiate to the seven cell types of the neural retina, the retinal pigmented epithelium and the optic stalk respectively. In anamniotes (fish, amphibia), the ciliary marginal zone (CMZ) of the neural retina contains a stem cell population that gives rise to all retinal cell types and facilitates life-long growth of the eye.
The lab is studying neuronal cell proliferation and differentiation in the developing, growing and regenerating eye and brain of fish (zebrafish, medaka) as model system. We are combining genetic, molecular and cell biological approaches with advanced imaging approaches to decipher the basic mechanisms that govern the balance of cell proliferation and differentiation in vivo. Special emphasis is given to follow the fate of proliferating and differentiating cells in the context of the fish retina and brain and to establish tools that allow visualizing the those processes in vivo. We take advantage of the life-long proliferation of retinal stem cells from the ciliary marginal zone (CMZ) that facilitates the continuous study of cells exiting the stem cell niche at the CMZ and their subsequent stereotypic differentiation.

We delineated factors involved in retinal growth and regeneration and the underlying transcriptional networks. A particular focus was on the role of retinal stem and progenitor cells and their role in establishing and maintaining the perfect shape of the eye which is fundamental for its functional homeostasis. We have shown that the neuroretina is creating shape out of itself via a programmed behavior of neuroretinal stem cells. The pigmented epithelium on the other side arises from the same stem cell niche and passively follows the lead (and shape) implemented by the action of the neuroretinal stem cells. Our computational model for retinal growth is complemented by functional insights originating from clonal gain and loss of function analyses of key players governing the activity of retinal stem and progenitor cells. We have employed stochastic activation of transcriptional modules that couple in vivo indicators with the gain or loss of function of key pathway components (e.g. of wnt signaling).

Our massive progress in Crispr/Cas technology was instrumental for the establishment of genetically validated conditional paradigms that now allow addressing the acute loss of key players (e.g. Rx genes). Those are of high interest since they appear to facilitate life-long growth of the retina in teleosts and their targeted inactivation has furthered our understanding of vertebrate retinal size control. Another striking feature of teleost eyes retinae is their apparently unlimited regenerative capacity. We carefully compared different species and took advantage of the loss of retinal regeneration in medaka (similar to human) to identify key factors that, when targeted to Mueller glia cells in the retina, reinstate the regenerative capacity.

Addressing the role of individual key genes in retinal development, growth and regeneration we initially focussed on the level of the population (tissue, organ). However, we soon realized that the function is only understood, when analysed on the level of the individual cell in the context of the population.

Scaling that insight, we had been initiating a large project already 2005 in collaboration with Ewan Birney (EBI) and have identified an unstructured medaka population in collaboration with K. Naruse from the NIBB in Okazakai, Japan in 2010. This was the starting point for systematic inbreeding in collaboration with Felix Loosli at KIT and we have now sequenced the genome more than 100 inbred lines (backcross generation 10 or higher, with E. Birney).

While the genetic resource was getting on its way, we in parallel established the resources for systematic quantitative analyses, with a focus on high resolution morphometrics, the heart, pharmacogenomics, stem cells and regeneration as well as behavioural analyses. We have established efficient and quantitative phenotyping pipelines adapted to high throughput.

Future research goals

In the coming years, we will merge the population genomics approaches and the studies on stem cells and regeneration to address their relevance in the context of environmental conditions. We will expand the panel in its width and will establish new and complementary inbred lines. In parallel we will integrate new features into the existing panel and will employ Crispr/Cas to engineer a unique and highly efficient PhiC31 landing site into a fully accessible locus (UBI-one) in each of the 120 inbred lines. This will facilitate the use of established assays in a fully comparable context, in the absence of different position effects either in the individual fish lines or in cell lines derived from them.

We will take advantage of the population genomics resource and establish further high throughput phenotyping pipelines to quantify regenerative and stem cell related phenotypes. One particular focus will be on the interplay between stem cells and the immune system, an unexpected and tight interconnection that we recently uncovered.

Another novel aspect of our work in the present years related to the medaka heart and the contribution of genetic and environmental factors to heart function at post-embryonic stages. GWAS on heart phenotypes is most advanced and we will use the heart as model for validating the relative contribution of individual SNPs to the heart phenotypes. We will first take advantage of newly established tools to map regulatory elements from human to medaka, in particular now incorporating the information provided by the sequenced genome of bridging species like the spotted gar.

Taken together our approaches will be directed towards ultimately allow addressing the relative contribution of multiple factors and will be a first step towards addressing the genetics of individuality in different environmental contexts.

Jochen Wittbrodt is a member of the CellNetworks Cluster of Excellence, the collaboartive research centres SFB873 on stem cells wnt the SFB1324 and the HBIGS graduate school. Our work is supported by funds from a variety of public sources, including the DFG, the BMBF and the European Commission via the European Research Council ERC.